The development of an effective AIDS vaccine remains one of the highest priorities in HIV-research. Because increasing evidence suggests that an effective vaccine will require both humoral and cellular immunity, this HIV-RAD project proposes to induce and measure both cellular and humoral immunity using plasmid DMAconstructs expressing SIV gag,-pol-env genes plus novel chemokine DNA plasmids to address mucosal cellular and humoral immunity. This proposal describes a new approach to inducing systemic and mucosal immunity. The overall objective of project #3 is to evaluate the potential of intramuscular (IM) delivery of chemokines to enhance mucosal immunity in the SIVmac model. The hypothesis to be tested is that chemokine-induced redirected trafficking of mucosal-homing immune cells to the systemic compartment leads to cellular and humoral mucosal immune responses without mucosal immunization. Mucosal-expressed chemokines CTACK, MECK, TECK will be tested for their individual potential to enhance the mucosal immune response of a systemically delivered SIV DNA vaccine. The vaccination approach will be IM co-immunization of macaques with an optimized SIVgag/pol/env plus each plasmid form of the mucosal-expressed chemokines. The specific aims are: 1 A. To measure antigen specific cell mediated immune responses systemically in blood as well as in the mucosa following intramuscular plasmid co-immunization of rhesus macaques with 3 individual chemokines plus pSIVgag/pol/env. This aim is expanded to test 5 monkeys per group compared to 3 per group in the previous submission. 1B. To measure antigen specific antibody mediated immune responses systemically in blood and lymph node tissues as well as in the mucosa following IM co-immunization of rhesus macaques with individual chemokines plus pSIVgag/pol/env. Samples from animals in 1Awill also be provided to the University of Alabama Birmingham (DAB) group headed by Dr. Jiri Mestecky. 2. To carry out animal efficacy studies using the SIVmac-rhesus model to evaluate a) 1 selected chemokine adjuvant, b) pSIVgag/pol/env plus the control plasmid and c) a 3rd group of 8 will receive control plasmids only. All animals will be inoculated by the rectal route with pathogenic SIVmac251. Materials will be provided to investigators at the TNPRC, University of Pennsylvania (UPenn) and UAB. This aim is expanded to 8 animals per group to increase the statistical power of the experiment.